Molecular analyses of T
plants from controlled crosses between MB17 and non-transgenic plants. (a) PCR for identification of transgene-excised progeny when MB17 was used as the male parent (WT × MB17 cross). 1–6, individual T1 plants. Lanes: e, PCR reactions for detection of autoexcision (primers e3&e4); b, reactions for the marker gene bar (primers b1&b2); r, reactions for the recombinase gene bxbNom (primers r3&r4). (b) PCR for determination of the segregation ratio in the T1 generation when MB17 was used as the female parent (MB17 × WT cross). 1–9, individual T1 plants. PCR reactions: same as reactions r and b in (a). (c) Southern blot hybridization for detection of the pMBXS824 T-DNA (Figure 3) in the genome of WT × MB17 progeny. Lanes: P, pMBXS824 DNA; WT, wild-type plant used as the female parent; T
, MB17 plant used as the pollen donor; a-f, individual T1 plants. Based on the results from PCR screening (Table 1), the analyzed T1 plants were categorized as follows: a-c, transgene-excised; d-e, transgenic; f, a non-transgenic T1 plant (a null segregant).